Substantial research has been undertaken in order to realize the possibility that the course of disease may be affected through the introduction of nucleic acids into living organisms. When utilized for medical purposes, such therapy, termed “gene therapy”, allows the alteration of the genetic repertoire of cells for a therapeutic benefit.
A variety of methods have been described for delivering nucleic acids to cells, including for example, the use of viral vectors derived from retrovirus, adenovirus, poxvirus, herpes virus, and adeno-associated virus (see Jolly, Cancer Gene Therapy 1:51-64, 1994), as well as direct nucleic acid transfer techniques, including direct DNA injection (Donnelly, J. J. et al., Ann. Rev. Immunol. 15:617-648, 1997), micro-projectile bombardment (Williams et al., PNAS 88:2726-2730, 1991), and nucleic acid complexes with lipids of several types (Felgner et al., Proc. Natl. Acad. Sci. USA 84:7413-7417, 1989). Many of these strategies are being applied widely, with clinical trials now ongoing for a wide range of hereditary (e.g., ADA deficiency, cystic fibrosis, hemophilia) and acquired (e.g., cancer, viral infection) diseases (Crystal, Science 270:404-410, 1995).
One particularly important application of gene therapy is in the vaccine field. Briefly, live virus vaccines (e.g., vaccinia, polio, measles) have been enormously successful and have made a dramatic and historic impact on public health. However, for certain pathogens, such as HIV, safety concerns about either reversion of attenuated vaccine strains to virulent phenotypes or inducing fulminate infection in immune compromised individuals have forced the development of subunit or inactivated virus vaccines. Unfortunately, in many cases, these vaccines have not elicited the potent broad-based humoral, cellular, and mucosal immune responses or long-term memory necessary to confer life-long protection in immunized individuals. Induction of such robust immune responses will be particularly important for vaccines against HIV and HCV infection, both of which have reached worldwide epidemic proportions and have, to date, proved elusive to candidate vaccines.
Still relatively new, plasmid DNA-based vaccines combine the latest developments of molecular biology with an ever-increasing understanding of immunology, and show promise for immunization against infection with pathogens, such as HIV, as well as other diseases for which improved vaccines are needed (e.g., influenza virus). Importantly, DNA immunization elicits humoral and cellular immune responses in rodents and non-human primates. DNA is an attractive mode for vaccination, as it provides the safety advantages of subunit or inactivated virus vaccines, and induces the MHC class I-restricted cytotoxic T cell responses typical of live virus, or “replicating antigen” vaccines, which may be critical for efficacy (Donnelly, J. J. et al., ibid).
Unfortunately however, recent DNA vaccine clinical trials have demonstrated that, although antigen-specific cytotoxic T lymphocyte responses were observed in vaccinated individuals, these responses were insufficient to afford protection against challenge with the infectious agent (Wang, R. et al., Science 282:476-480, 1998; Calarota, S. et al., Lancet 351:1320-1325, 1998). Thus, for DNA-based vaccination to become a widely used method for protection against infectious disease and cancer, the technology must be further improved.
The present invention provides compositions and methods for enhancing the efficacy of DNA vectors, as well as a variety of other gene delivery vectors, used for genetic immunization. Such improvements can include immunostimulatory modifications to the antigen-expressing vector itself, or the in vivo administration, along with the antigen-expressing vector, of one or more additional gene delivery vectors that are immunostimulatory. The improvements described herein address previous difficulties associated with the use of gene delivery vectors by enhancing the extent of the antigen-specific immune response, thus providing the broad and robust responses critical for successful prophylactic or therapeutic vaccination against infectious agents and cancer.